In analytical chemistry, liquid chromatography (LC) and gas chromatography (GC) techniques have become important tools in the identification of chemical sample components. The basic principle underlying all chromatographic techniques is the separation of a sample chemical mixture into individual components by transporting the mixture in a carrier fluid through a porous retentive media. The carrier fluid is referred to as the mobile phase and the retentive media is referred to as the stationary phase. The principal difference between liquid and gas chromatography is that the mobile phase is either a liquid or a gas, respectively.
In a GC apparatus, an inert carrier gas typically is passed through a temperature-controlled column which contains a stationary phase in the form of porous sorptive media. Gas chromatography columns have also been known to comprise a hollow capillary tube having an inner diameter in the range of few hundred microns coated with the stationary phase. A sample of the subject mixture is injected into the carrier gas stream and passed through the column. As the subject mixture passes through the column, it separates into its various components. Separation is due primarily to differences in the partial pressures of each sample component in the stationary phase versus the mobile phase. These differences are a function of the temperature within the column. A detector, positioned at the outlet end of the column, detects each of the separated components contained in the carrier fluid as they exit the column.
The analytical choice between liquid and gas chromatography techniques is largely dependent on the molecular weight of the components to be analyzed. Liquid chromatography devices are capable of analyzing much heavier compounds than gas chromatography devices. However, gas chromatography detection techniques are more sensitive and therefore are generally preferred.
The advent of supercritical fluid chromatography (SFC) provided a potential bridge between gas and liquid chromatography by providing relatively high sensitivity for higher molecular weight samples. In SFC, a fluid heated above its critical point is used as the mobile phase. This fluid is passed under pressure through a media which differentially retains sample components. As the pressure of the mobile phase is increased, for example, from about 40 atmospheres to approximately 400 atmospheres, the sample being analyzed separates into its various components in relation to the relative differential solubility of each component in the mobile phase. Since the mobile phase is a gas, the same detectors used in GC techniques can be utilized, significantly enhancing detector sensitivity and selectivity.
SFC has been found to be primarily useful in the analysis of compounds having molecular weights in the range of about 100 to about 10,000 and in the analysis of thermally labile molecules such as pesticides and pharmaceuticals. The problem with SFC, however, is that a considerable amount of time is required to conduct a sample analysis.
One approach to improved chromatographic devices has focused on programmed computer control. It is known, for example, to program the column temperature of a GC device. Since separation of the sample components is due primarily to differences in their partial pressures in the stationary phase versus the mobile phase and since these differences are a function of the temperature within the column, raising the column temperature either in a constant linear fashion or in a stepwise linear fashion over a sufficient range of temperature can assure high resolution of each sample component in a minimized time period.
It is also known that the time required for a temperature programmed GC analysis can be reduced even further by programming the flow rate of the carrier gas. Scott, R. P. W., "New Horizons in Column Performance", Gas Chromatography, 1964, 32-37 indicates that analysis time can be reduced by increasing the flow rate. However, while increasing the flow rate may reduce analysis time, efficiency and resolution are also reduced. This reduction may be accepted for analytes that have excess resolution on the column. This is similar to increasing the column temperature, which may not be possible for particularly labile analytes or stationary phases.
Costa Neto, C., et al., Journal of Chromatography, 1964, 15, 301-313 discusses programming the flow of the GC mobile phase in isothermic or temperature programmed runs in order to obtain the separation of complex mixtures. Costa Neto, et al. discuss the theoretical derivation of equations which relate flow rate to various chromatogram properties such as peak migration, peak width, peak area and peak height. Certain derivations also relate flow rate to efficiency and resolution. The programmed flow actually used by the authors was said to be manual in nature using a step valve.
Zlatkis, A., Journal of Gas Chromatography, March 1965, 75-81 discusses the use of a pneumatic flow controller for regulating flow rate in an exponential fashion between preset limits. In reviewing previous flow programming reports, such as the Costa Neto, et al. reference discussed above, Zlatkis et al. characterize that reference as discussing flow programming only in relation to so-called preparative gas chromatography, not practical analytical gas chromatography.
Nygren, S. et al., Journal of Chromatography, 1976, 123, 101-108 discuss flow programming through the use of a metering valve in the side outlet of an inlet splitter. Nygren, et al. state that results comparable to temperature programming could be achieved under certain circumstances by exponentially programming carrier gas flow.
More recently, Larson, J. R. et al., Journal of Chromatography; 1987, 405, 163-168 discuss a continuous flow programming technique for process capillary gas chromatography. However, these techniques do not have temperature programming capabilities. The authors concluded that by programming carrier gas flow in a process GC application, shorter cycle times could be achieved than with temperature programmed GC devices.
One problem with each of the flow programming devices discussed above is that carrier gas flow and/or column temperature systems operate independently of one another. In general, such independently operated closed loop systems are incapable of detecting undesirable conditions affecting the accuracy of the chromatographic analysis or of optimizing flow and temperature conditions simultaneously. Moreover, the systems are incapable of making adjustments to maintain multiple parameters at optimized conditions.
Thus, the invention disclosed by U.S. patent application Ser. No. 349,740, now U.S. Pat. No. 4,994,096, in the names of Klein, et al. represented a significant advance in the programming of chromatographic flow. Application Ser. No. 349,740, now U.S. Pat. No. 4,994,096 which is incorporated herein by reference, disclosed an open loop system for controlling the flow rate of a carrier fluid in a system wherein a portion of the chromatographic column is subjected to a temperature profile. By this open loop system, Klein, et al. overcame many of the problems of prior temperature and flow programming devices and also extended the molecular weight range of compounds capable of GC analysis.
However, the flow and temperature programming disclosed in application Ser. No. 349,740, now U.S. Pat. No. 4,994,096, is primarily directed toward effective compound chromatographic resolution. Neither that application nor the prior art focus on improving the detection of the resolved components. In fact, due to manner in which the flow rate of carrier fluids exiting a temperature-programmed column will vary over time, such fluids are not well suited for introduction into many of the flow-sensitive or pressure-sensitive chromatographic detectors commonly employed in the art. Variations in the flow rates or pressures of carrier fluids introduced into such detectors frequently will result in variability and non-optimization of detector response. There thus exists a need for a chromatographic device wherein the flow rate of the carrier fluid entering the chromatographic detector is carefully controlled to provide optimal detector response.
In addition, many of the commonly-employed chromatographic detectors require the provision of one or more support fluids. For example, the operation of detectors such as the flame photometric detector depends intimately upon the composition, pressure, and/or flow rate of several flame support fluids. Detector response attributes such as sensitivity, specificity, linear dynamic range, stability, and noninterference are also known to be dependent upon each support fluid and its physicochemical properties. However, none of the chromatographic devices presently known in the art are able to assure controlled provision of these support fluids. By carefully controlling the provision of detector support fluids, it may be possible to optimize one or more of the detector response attributes over the course of a chromatographic run.